title: Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of SpyCEP, a candidate antigen for a vaccine against Streptococcus pyogenes.
abstract: Streptococcus pyogenes (Group A streptococcus; GAS) is an important human pathogen against which an effective vaccine does not yet exist. The S. pyogenes protein SpyCEP (S. pyogenes cell-envelope proteinase) is a surface-exposed subtilisin-like serine protease of 1647 amino acids. In addition to its auto-protease activity, SpyCEP is capable of cleaving interleukin 8 and related chemokines, contributing to GAS immune-evasion strategies. SpyCEP is immunogenic and confers protection in animal models of GAS infections. In order to structurally characterize this promising vaccine candidate, several SpyCEP protein-expression constructs were designed, cloned, produced in Escherichia coli, purified by affinity chromatography and subjected to crystallization trials. Crystals of a selenomethionyl form of a near-full-length SpyCEP ectodomain were obtained. The crystals diffracted X-rays to 3.3 Å resolution and belonged to space group C2, with unit-cell parameters a=139.2, b=120.4, c=104.3 Å, β=111°.
authors.affiliation: Research Centre, Novartis Vaccines and Diagnostics, Via Fiorentina 1, 53100 Siena, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Abate
authors.forename: Enrico
authors.initials: E
authors.lastname: Malito
authors.forename: Fabiana
authors.initials: F
authors.lastname: Falugi
authors.forename: Immaculada
authors.initials: I
authors.lastname: Margarit Y Ros
authors.forename: Matthew James
authors.initials: MJ
authors.lastname: Bottomley
mesh_terms: D000942:Antigens, Bacterial / Q000737:chemistry* / Q000302:isolation & purification*; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D006801:Humans; D010447:Peptide Hydrolases / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*; D017434:Protein Structure, Tertiary; D012645:Selenomethionine / Q000737:chemistry; D022541:Streptococcal Vaccines / Q000276:immunology*; D013297:Streptococcus pyogenes / Q000276:immunology*; D014961:X-Ray Diffraction
chemical_list: D000942:Antigens, Bacterial; D022541:Streptococcal Vaccines; D012645:Selenomethionine; D010447:Peptide Hydrolases; C527349:SpyCEP protein, Streptococcus pyogenes
keywords: GAS; SpyCEP; Streptococcus pyogenes; auto-proteases; vaccine antigens